title: The nature of the carbohydrate binding module determines the catalytic efficiency of xylanase Z of Clostridium thermocellum.
abstract: Xylanase Z of Clostridium thermocellum exists as a complex in the cellulosome with N-terminus feruloyl esterase, a carbohydrate binding module (CBM6) and a dockerin domain. To study the role of the binding modules on the activity of XynZ, different variants with the CBM6 attached to the catalytic domain at its C-terminal (XynZ-CB) and N-terminal (XynZ-BC), and the CBM22 attached at N-terminus (XynZ-B'C) were expressed in Escherichia coli at levels around 30% of the total cell proteins. The activities of XynZ-BC, XynZ-CB and XynZ-B'C were 4200, 4180 and 20,700U μM(-1) against birchwood xylan, respectively. Substrate binding studies showed that in case of XynZ-BC and XynZ-CB the substrate birchwood xylan remaining unbound were 51 and 52%, respectively, whereas in the case of XynZ-B'C the substrate remaining unbound was 39% under the assay conditions used. The molecular docking studies showed that the binding site of CBM22 in XynZ-B'C is more exposed and thus available for substrate binding as compared to the tunnel shape binding pocket produced in XynZ-BC and thus hindering the substrate binding. The substrate binding data for the two constructs are in agreement with this explanation.
authors.affiliation: School of Biological Sciences, University of the Punjab, Lahore, Pakistan.
authors.forename: Muhammad Imran M
authors.initials: MI
authors.lastname: Khan
authors.forename: Muhammad
authors.initials: M
authors.lastname: Sajjad
authors.forename: Saima
authors.initials: S
authors.lastname: Sadaf
authors.forename: Rehan
authors.initials: R
authors.lastname: Zafar
authors.forename: Umer H K
authors.initials: UH
authors.lastname: Niazi
authors.forename: Muhammad Waheed
authors.initials: MW
authors.lastname: Akhtar
mesh_terms: D001665:Binding Sites; D002241:Carbohydrates / Q000737:chemistry*; D002384:Catalysis; D020134:Catalytic Domain; D003001:Cloning, Molecular; D048013:Acetivibrio thermocellus / Q000201:enzymology* / Q000254:growth & development; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D015964:Gene Expression Regulation, Bacterial; D062105:Molecular Docking Simulation; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
chemical_list: D002241:Carbohydrates; D043364:Endo-1,4-beta Xylanases
keywords: Activity enhancement; C. thermocellum; Carbohydrate binding module; Thermostability; Xylanase Z